| dc.description.abstract | The fibroblast growth factor (FGF) signalling pathway contributes to the regulation of a variety of cellular functions, affecting differentiation, migration, proliferation, and survival. Unsurprisingly, cancer cells can hijack this pathway for growth or survival advantages, through alterations in ligands, receptors or regulatory molecules. Sequencing consortia have highlighted how mutation, amplification, translocation or loss of elements in the FGF signalling network can contribute to tumourigenesis, and the pathway is a major clinical target. Many FGF receptor (FGFR) driven cancers develop resistance against commonly used receptor tyrosine kinase (RTK) targeted therapeutics and dissection of the mechanisms that underlie this, both in the cancer cell, and also via stromal crosstalk in the tumour microenvironment, is of utmost importance to the development of therapeutic approaches to treat FGFR-driven cancers. In this work, I focus on the mechanisms of FGF deregulation and the impact of stromal cells. I aim to identify the implications of FGFR2 aberrations on gastric and endometrial cancer, and FGFR1 aberrations on lung cancer. Furthermore, I aim to dissect the mechanisms by which targeted cells may develop drug resistance both in two-dimensional (2D) and in a more physiomimetic three-dimensional (3D) co-culture model. It was established that cancer cells with FGFR aberrations were sensitive towards FGFR inhibitors such as PD173074 (PD), AZD4547 (AZD) and BGJ398 (BGJ). Cancer cells could be killed with increasing concentration of these drugs and exhibited sensitivity to FGFR inhibitors by decreased p-AKT and p-ERK signalling. However, with prolonged exposure to FGFR inhibition, certain cells began to acquire resistance in 2D. To study drug resistance in a more physiomimetic environment, cells were grown alone or co-cultured with fibroblasts and treated with FGFR inhibitors PD, AZD or BGJ and imaged using fluorescent live cell imaging. These cultures were then fixed, paraffin embedded and immuno- or Haematoxylin and Eosin (H&E) stained. To study the emergence of drug resistance in 3D with and without stromal support, cells were seeded into Alvetex® scaffolds and treated with increasing concentrations of BGJ until resistant populations were observed via confocal fluorescence microscopy. Cancer cells grown in co-culture with fibroblasts acquired resistance faster than monoculture cells. Significantly regulated genes between resistant and parental mono- and co-culture cells were identified using ribonucleic acid sequencing (RNA-Seq) and bioinformatics. The majority of significantly regulated pathways in FGFR2-amplifed gastric cancer cells, were metabolic pathways including retinol metabolism, starch and sucrose degradation, which was mainly driven by sucrose-isomaltase (SI) and aldolase B (ALDOB), resulting in glucose generation. Targets were then modified using different approaches such as knockdown, overexpression and drug treatments to observe how this affects resistance of FGFR-driven cancers. Blockade of ALDOB resulted in lower cell numbers within 2D cultures in drug-resistant gastric cancer cells, possibly by slowing down cell growth rather than inducing cell death. However, the effect of glucose or sucrose on cells is possibly through a SI-independent manner. When analysing gene expression in cells grown on 2D versus 3D substrates, expression levels varied considerably, and this underlines the importance of investigating biological processes in a physiomimetic in vivo-like setting. Drug resistance in cancer is an increasing issue for successful therapy and a number of interesting targets have been identified in FGFR2-amplified gastric cancer that could be responsible for drug resistance towards FGFR inhibitors. With stromal support, cancer cells acquired resistance faster, which highlights the influence of fibroblasts and the extracellular matrix (ECM) environment on resistance. It was shown that sugar metabolism plays a substantial part in resistance, together with the retinol pathway. Possibly, these genes and pathways are intertwined, ultimately aiding cancer cells to grow in presence of inhibition via cytoprotective mechanisms and also generation of metabolites to ensure sustained proliferation of cells. The importance of these genes and pathways need to be further evaluated. Therefore, drug combination therapy could be the pivotal way forward to hinder cancer cells from rewiring their pathways to overcome drug inhibition. | en_US |
